Multiple glacial refugia and contemporary dispersal shape the genetic structure of an endemic amphibian from the Pyrenees.

Historical factors (colonization scenarios, demographic oscillations) and contemporary processes (population connectivity, current population size) largely contribute to shaping species' present-day genetic diversity and structure. In this study, we use a combination of mitochondrial and nuclear DNA markers to understand the role of Quaternary climatic oscillations and present-day gene flow dynamics in determining the genetic diversity and structure of the newt Calotriton asper (Al. Dugès, 1852), endemic to the Pyrenees. Mitochondrial DNA did not show a clear phylogeographic pattern and presented low levels of variation. In contrast, microsatellites revealed five major genetic lineages with admixture patterns at their boundaries. Approximate Bayesian computation analyses and linear models indicated that the five lineages likely underwent separate evolutionary histories and can be tracked back to distinct glacial refugia. Lineage differentiation started around the Last Glacial Maximum at three focal areas (western, central and eastern Pyrenees) and extended through the end of the Last Glacial Period in the central Pyrenees, where it led to the formation of two more lineages. Our data revealed no evidence of recent dispersal between lineages, whereas borders likely represent zones of secondary contact following expansion from multiple refugia. Finally, we did not find genetic evidence of sex-biased dispersal. This work highlights the importance of integrating past evolutionary processes and present-day gene flow and dispersal dynamics, together with multilocus approaches, to gain insights into what shaped the current genetic attributes of amphibians living in montane habitats.

Acceleration of cyanobacterial dominance in north temperate-subarctic lakes during the Anthropocene.

Increases in atmospheric temperature and nutrients from land are thought to be promoting the expansion of harmful cyanobacteria in lakes worldwide, yet to date there has been no quantitative synthesis of long-term trends. To test whether cyanobacteria have increased in abundance over the past ~ 200 years and evaluate the relative influence of potential causal mechanisms, we synthesised 108 highly resolved sedimentary time series and 18 decadal-scale monitoring records from north temperate-subarctic lakes. We demonstrate that: (1) cyanobacteria have increased significantly since c. 1800 ce, (2) they have increased disproportionately relative to other phytoplankton, and (3) cyanobacteria increased more rapidly post c. 1945 ce. Variation among lakes in the rates of increase was explained best by nutrient concentration (phosphorus and nitrogen), and temperature was of secondary importance. Although cyanobacterial biomass has declined in some managed lakes with reduced nutrient influx, the larger spatio-temporal scale of sedimentary records show continued increases in cyanobacteria throughout the north temperate-subarctic regions.

Ice phenology interactions with water and air temperatures in high mountain lakes.

Ice phenology is of great importance for the thermal structure of lakes and ponds and the biology of lake species. Under the current climate change conditions, ice-cover duration has been reduced by an advance in ice-off, and a delay in ice-on, and future projections foresee this trend as continuing. Here, we describe the current ice phenology of Pyrenean high mountain lakes and ponds, including ice-cover duration and ice-on and ice-off dates. We used mixed models to identify the variables that explained the observed patterns, extrapolated them across all water bodies in the mountain range, and related the seasonality of air and water temperatures with ice phenology using structural equation models. Ice phenology was obtained from the temperature series of 85 lakes and ponds for fourteen years, including 2001 to 2004 and 2009 to 2019. We discovered that high autumn precipitation was related to earlier ice-on dates, and that earlier ice-off dates were associated with higher following-summer water temperatures. We found a greater predictability of ice-off dates and ice-cover duration than ice-on dates. Altitude was the most important variable explaining the variation in ice phenology, followed by latitude, which was related to climatic differences among the northern and southern slopes of the mountain range. The lake area was significant for ice-on dates and ice-cover duration. The interannual variability in air temperature and radiation was remarkable for the ice-off date and ice-cover duration but not for the ice-on date. In contrast, wind speed was related to an earlier ice-off date and shorter ice-cover duration. All the measured lakes and ponds froze in winter during the studied period, a feature maintained in the extrapolation to the whole set of water bodies.

Simple on-site extraction and GC-MS analysis of rotenone and degradation products for monitoring invasive fish eradication treatments in fresh and brackish waters.

The introduction of invasive fish species to aquatic ecosystems has been demonstrated to cause disastrous ecological effects. Current conservation strategies regard rotenone-containing piscicide formulations, such as commercial product CFT Legumine, as a potentially viable alternative to the cumbersome traditional approaches to fish eradication. This consideration relies on the fast degradation of rotenone and its relatively rapid dissipation from the environment. Piscicide treatments in fragile aquatic ecosystems should thus monitor not only rotenone concentrations following application, but also other byproducts and degradation products. We present a methodology for the analysis of rotenoids in fresh and brackish waters that addresses two main challenges: the accurate determination of applied concentrations in different salinity concentrations by performing a simplified on-site solid-phase extraction, overcoming the fast degradation of rotenone in sample storage conditions, and the selective analysis of rotenoid byproducts and degradation products by gas chromatography coupled to mass spectrometry. Limits of quantification were below the ecological no-effect concentration of rotenone (2 µg/L) and average recoveries exceeded 80%. Accuracy (compared to expected values) and precision (deviation of replicates) ranged from 78 to 103% and 3 to 14%, respectively, across various rotenoid concentrations. These metrics are more than satisfactory for the intended application of this simplified procedure. The method was applied to piscicide-treated samples, revealing significant and fast degradation of parent rotenoids in storage conditions, as well as a non-negligible accumulation of rotenone in the particulate fraction of water that could impact the effectivity of eradication efforts.

Factors of surface thermal variation in high-mountain lakes of the Pyrenees.

Thermal variables are crucial drivers of biological processes in lakes and ponds. In the current context of climate change, determining which factors better constrain their variation within lake districts become of paramount importance for understanding species distribution and their conservation. In this study, we describe the regional and short-term interannual variability in surface water temperature of high mountain lakes and ponds of the Pyrenees. And, we use mixed regression models to identify key environmental factors and to infer mean and maximum summer temperature, accumulated degree-days, diel temperature ranges and three-days' oscillation. The study is based on 59 lake-temperature series measured from 2001 to 2014. We found that altitude was the primary explicative factor for accumulated degree-days and mean and maximum temperature. In contrast, lake area showed the most relevant effect on the diel temperature range and temperature oscillations, although diel temperature range was also found to decline with altitude. Furthermore, the morphology of the catchment significantly affected accumulated degree-days and maximum and mean water temperatures. The statistical models developed here were applied to upscale spatially the current thermic conditions across the whole set of lakes and ponds of the Pyrenees.

Factors regulating primary producers' assemblages in Posidonia oceanica (L.) Delile ecosystems over the past 1800 years.

Posidonia oceanica (L.) Delile meadows are highly productive coastal marine ecosystems that provide multiple ecosystem services. The seagrass is not always the major contributor to total primary production, however, little is known about long-term changes in the composition of primary producers within seagrass meadows. Understanding compositional shifts within the community of primary producers is crucial to evaluate how climate and anthropogenic change affect the functioning of seagrass ecosystems. Here we analysed marker pigment composition in seagrass cores from two bays of the Cabrera Island (Balearic Islands, Spain) to asses long-term changes in phototrophic community composition and production in seagrass meadows, and identify the environmental factors triggering those changes. The proxy dataset was explored using principal component analyses (PCA): one including the pigment dataset to look for associations between producers' groups, and another one combining the pigment dataset with plausible local and global regulatory factors to assess the environmental drivers of change. Analyses of characteristic pigments and morphological fossils (cysts) showed that the abundance of dinoflagellates increased over the last 150-300 years, coeval with a rise in solar irradiance and air temperature. When compared among embayments, pigments from cyanobacteria predominated in seagrass meadows located at Es Port, a sheltered bay receiving higher terrestrial runoff; whereas pigments from diatoms, seagrasses and rodophytes were more common at Santa Maria, an exposed bay with clearer waters. Water depth also played a role in controlling the phototrophic community composition, with greater abundance of diatoms in the shallowest waters (<5 m). Overall, our results suggested that historical and spatial variation in seagrass meadows' phototrophic community composition was influenced by the interaction between local factors (catchment-bay characteristics) and global climate processes (energy influx). Together these patterns forecast how marine primary producers and seagrass ecosystem structure may respond to future global warming.

Responses of fish assemblage structure to large-scale weir construction in riverine ecosystems.

Worldwide, increasing amounts of dams and weirs have been established in rivers in recent decades, often with drastic effects on their ecosystems. Between late 2009 and 2011, 16 large-scale dams were built in the main channels of the four largest rivers in South Korea, eight of these along the main channel of Nakdong River (300 km, 520 km in total). We studied the effect of these constructions on the fish community in the riparian zone based on intensive fish field surveys conducted in the littoral zone during 2007-2017, analysis of fishermen's catch data and molecular analysis of the diet of the keystone species, largemouth bass (Micropterus salmoides). Our results, based on RDA and GLM analyses, showed that environmental characteristics and fish species composition changed significantly with dam construction. Total fish abundance and biomass decreased after the start of the weir construction and fish richness decreased with time. The abundance and biomass of exotic fish decreased during construction but recovered afterwards, leading to higher proportions of exotics. Dominance and Shannon indices did not change significantly from before to after construction, while the Evenness index showed a significant decreasing trend. A comparison of the diet composition of largemouth bass showed decreasing genetic variation after construction. The changes in Functional Feeding Group (FFG) of the benthic communities at the study sites did not coincide with FFG changes in the prey items of largemouth bass, indicating a transition in feeding mode from before to after construction. In conclusion, the endemic and native fish species were most sensitive and showed lower resilience to disturbance by the large-scale dam construction than the translocated and exotic species, and the lake-like ecosystems after construction markedly improved the competitive capacity of these exotic fish over the native and endemic species in the riparian zone of the river.

Non-native Minnows Threaten Quillwort Populations in High Mountain Shallow Lakes.

Submersed aquatic plants are a key component of shallow, clear water lakes contributing to primary production and water quality. High mountain lakes are naturally fishless although invasive trout and most recently minnows have been introduced causing a major impact on fauna richness. The Pyrenean high mountain range has preserved soft-water oligotrophic boreal isoetids in their southern limit of distribution but the recent fish introduction is a potential factor of stress that needs to be addressed. We here work under the hypothesis that due to contrasting ecological features, trout will not be heavily affecting quillwort populations while minnows will have a stronger effect on zooplankton and zoobenthos that will promote algal growth and reduce light availability for the underwater meadows. Ten Pyrenean shallow lakes representative of three scenarios -fishless, with trout and with minnows-, were sampled for meadow structure, water column and benthic environment characterization in mid-summer 2015 and 2016. Quillwort biomass allocation (above vs. belowground), epiphytic load, and composition of the algal community (abundant cyanobacteria) differed in the presence of minnows. In trout lakes biomass allocation and epiphytic load were average and the algal community composed by chlorophytes and diatoms as in fishless lakes. Biomass ratio was close to thresholds of negative buoyancy in minnow lakes indicating that meadows were at risk of uprooting and consequent de-vegetation. Total and soluble carbohydrates were lower and the sporangia contained significantly less reserves to constrain growth and expansion in the presence of minnows. Lake scenarios were coupled to physicochemical differences with low light, high phosphorus and Chl-a (mesotrophia) in minnow lakes, while trout and fishless lakes remained oligotrophic. This is the first study assessing the impact of non-native fish on soft-water isoetids from mountain lakes and shows that minnows are a major threat to quillworts. The impaired light environment (from epiphytic algal overgrow and water column Chl-a) entails consequent regression (i.e., no recruitment) and de-vegetation (uprooting) of the meadows. Since soft-water oligotrophic mountain lakes are protected under the Habitats Directive, some action needs to be urgently implemented not only to preserve quillworts but to the overall ecological integrity of the lakes.

Historical shifts in oxygenation regime as recorded in the laminated sediments of lake Montcortès (Central Pyrenees) support hypoxia as a continental-scale phenomenon.

Recent expansion of anoxia has become a global issue and there is potential for worsening under global warming. At the same time, obtaining proper long-term instrumental oxygen records is difficult, thus reducing the possibility of recording long-term changes in oxygen shifts that can be related with climate or human influence. Varved lake sediments provide the better time frame to study this phenomenon at high resolution. We tracked the oxic/anoxic shifts of the varved Lake Montcortès since 1500CE, and tried to recognise anthropogenic and climatic influences combining biological and geochemical proxies. Four main scenarios emerged: 1) years with abrupt sediment inputs (A); 2) years with outstanding mixing and oxygenation of the water column (B); 3) years with strong stratification, anoxia, intense sulfur bacterial activity and increased biomass production (C); 4) years with stratification and anoxia, but relatively less biomass production (D). In line with current limnologic trends, high supra-annual variability in the occurrence of oxygenation events was observed. Interestingly, at least 45.3% of the years were mixing years and, like the meromictic ones, were mostly clustered into groups of consecutive years, thus alternating years of monomixis with years of meromixis. Most years of D belong to the period 1500-1820CE, when human activities were the most intense. Most years of A belonged to the climatic unstable period of 1850-1899CE. Years of B were irregularly distributed but were best represented in the period 1820-1849CE. Most years of C belonged to the 20th century. More than 90% of the years with climatic instrumental records belonged to B and C. Current climate warming seems to be taking control over the oxygenation capacity of the lake, especially since the second half of the 20th century. Our results support recent findings related to hypoxia spreading at the global scale.

Discovering hidden biodiversity: the use of complementary monitoring of fish diet based on DNA barcoding in freshwater ecosystems.

Ecological monitoring contributes to the understanding of complex ecosystem functions. The diets of fish reflect the surrounding environment and habitats and may, therefore, act as useful integrating indicators of environmental status. It is, however, often difficult to visually identify items in gut contents to species level due to digestion of soft-bodied prey beyond visual recognition, but new tools rendering this possible are now becoming available. We used a molecular approach to determine the species identities of consumed diet items of an introduced generalist feeder, brown trout (Salmo trutta), in 10 Tasmanian lakes and compared the results with those obtained from visual quantification of stomach contents. We obtained 44 unique taxa (OTUs) belonging to five phyla, including seven classes, using the barcode of life approach from cytochrome oxidase I (COI). Compared with visual quantification, DNA analysis showed greater accuracy, yielding a 1.4-fold higher number of OTUs. Rarefaction curve analysis showed saturation of visually inspected taxa, while the curves from the DNA barcode did not saturate. The OTUs with the highest proportions of haplotypes were the families of terrestrial insects Formicidae, Chrysomelidae, and Torbidae and the freshwater Chironomidae. Haplotype occurrence per lake was negatively correlated with lake depth and transparency. Nearly all haplotypes were only found in one fish gut from a single lake. Our results indicate that DNA barcoding of fish diets is a useful and complementary method for discovering hidden biodiversity.